High cortisol levels can lead to a variety of different health issues and cortisol levels can increase in the aging population or related to stress. In many instances, the key to controlling intracellular levels of these molecules is their hydrolysis by cyclic nucleotide phosphodiesterases (PDEs). Cyclic nucleotide phosphodiesterases (PDEs) catalyze the hydrolysis of cyclic nucleotides, such as the second messengers cAMP (cyclic adenosine 3′5′-monophosphate) and cGMP (cyclic guanine 3′5′-monophosphate). Thus, different PDEs play pivotal regulatory roles in a wide variety of signal transduction pathways (Beavo, Physiol. Rev. 75: 725-48, 1995). For example, certain PDEs mediate processes involved in vision (McLaughlin et al., Nat. Genet. 4: 130-34, 1993), olfaction (Yan et al., Proc. Natl. Acad. Sci. USA 92: 9677-81, 1995), platelet aggregation (Dickinson et al. Biochem. J. 323: 371-77, 1997), aldosterone synthesis (MacFarland et al., J. Biol. Chem. 266: 136-42, 1991), insulin secretion (Zhao et al., J. Clin. Invest. 102: 869-73, 1998), T cell activation (Li et al., Science 283: 848-51, 1999), and smooth muscle relaxation (Boolell et al., Int. J. Impot. Res. 8: 47-52, 1996; Ballard et al., J. Urol. 159: 2164-71, 1998).
Mammals express eleven families of PDEs (3 cAMP-specific, 3 cGMP-specific, and 5 dual-specificity), encoded by 21 genes that produce over 100 distinct PDE isoenzymes due to alternative splicing of the transcript (Beavo, Physiol. Rev. 75: 725-48, 1995; Beavo et al., Mol. Pharmacol. 46: 399-05, 1994; Soderling et al., Proc. Natl. Acad. Sci. USA 95: 8991-96, 1998; Fisher et al., Biochem. Biophys. Res. Commun. 246: 570-77, 1998; Hayashi et al., Biochem. Biophys. Res. Commun. 250: 751-56, 1998; Soderling et al., J. Biol. Chem. 273: 15553-58, 1998; Fisher et al., J. Biol. Chem. 273: 15559-64, 1998; Soderling et al., Proc. Natl. Acad. Sci. USA 96: 7071-76, 1999; and Fawcett et al., Proc. Natl. Acad. Sci. USA 97: 3702-07, 2000). These enzymes perform distinct functions in the body due to tissue-specific expression, as well as subcellular localization of PDE isoforms and the effector proteins regulated by cAMP or cGMP levels. Given the complexity of this superfamily for which there are only two substrates; cAMP and cGMP, compounds that specifically and selectively inhibit individual PDEs can be powerful tools to advance our understanding of the function of a given PDE and can allow one to investigate the potential of that PDE as a therapeutic target.
However, given the difficulty of predicting transcriptional start sites and splice variants from primary genomic sequence data, it is still not known with exact certainty for any species how many different PDE mRNAs are transcribed. Furthermore, it also is not yet clear whether all transcript variants are present in all species.
Each PDE family is distinguished functionally by unique enzymatic characteristics and pharmacological profiles. In addition, each family exhibits distinct tissue, cell, and subcellular expression patterns (Beavo et al., Mol. Pharmacol. 46: 399-405, 1994; Soderling et al., Proc. Natl. Acad. Sci. USA 95: 8991-96, 1998; Fisher et al., Biochem. Biophys. Res. Commun. 246: 570-77, 1998; Hayashi et al., Biochem. Biophys. Res. Commun. 250: 751-56, 1998; Soderling et al., J. Biol. Chem. 273: 15553-58, 1998; Fisher et al., J. Biol. Chem. 273: 15559-64, 1998; Soderling et al., Proc. Natl. Acad. Sci. USA 96: 7071-76, 1999; Fawcett et al., Proc. Natl. Acad. Sci. USA 97: 3702-07, 2000; Boolell et al., Int. J. Impot. Res. 8: 47-52, 1996; Ballard et al., J. Urol. 159: 2164-71, 1998; Houslay, Semin. Cell Dev. Biol. 9: 161-67, 1998; and Torphy et al., Pulm. Pharmacol. Ther. 12: 131-35, 1999). Therefore, by administering a compound that selectively regulates the activity of one family or subfamily of PDE enzymes, it is possible to regulate cAMP and/or cGMP signal transduction pathways in a cell- or tissue-specific manner.
PDE11 is one of the most recently described families of PDEs; PDE11A has been identified (Fawcett et al., Proc. Natl. Acad. Sci. USA 97: 3702-07, 2000, hereinafter “Fawcett, 2000,” Yuasa et al., J. Biol. Chem. 275: 31469-79, 2000, hereinafter “Yuasa, 2000”). While PDE11A is known to be expressed in, e.g., testis, skeletal muscle, kidney, liver, various glandular tissue (e.g., pituitary, salivary, adrenal, mammary, and thyroid), pancreas, spinal cord, and trachea (Fawcett, 2000), little is known about PDE11A function. The present invention provides biological tools to study PDE11A function and methods to identify agents that regulate PDE11A activity for use in treating diseases and conditions that are linked to these PDE11A functions.
Presently, little is known about the PDE11 enzymes beyond their biochemical characteristics and basic genetics. Four variants of PDE11A have been identified (PDE11A1-4). The longest variant, PDE11A4, has two N-terminal GAF domains, whereas the other variants are truncations of this variant of varying lengths. The drug tadalafil (an approved PDE5 inhibitor) is also a potent inhibitor of PDE11, thus there is significant evidence that pharmacological inhibition of PDE11 is not harmful to humans.
It is clear that the PDE11A variants demonstrate differential tissue expression. In humans, PDE11A1 mRNA is most prominent in skeletal muscle and prostate. PDE11A3 mRNA is found specifically in testis and PDE11A4 mRNA is highly expressed in prostate. PDE11A protein localization studies have been somewhat contradictory in their findings, probably because of differences in the specificity of the antibodies used. PDE11A1 protein was originally detected in prostate and skeletal muscle, although a later study did not detect PDE11A1 protein in any tissues. In fact, only PDE11A4 protein has been verified and is found in prostate, pituitary, heart, and liver. Another study suggested that PDE11A is widely expressed, and immunohistochemistry using an antibody reported to recognize all PDE11A variants localized it to the epithelial, endothelial, and smooth muscle cells of many tissues, but at highest levels in the prostate, testis, kidney, adrenal gland, colon, and skin. However, a separate study did not find any PDE11 protein expression in human testis. As with many PDEs, it is still not clear if the same tissue, cellular, and subcellular localization is found among species. PDE11 is highly expressed in the testis, prostate, and developing spermatozoa.
Relatively little is known about the function of PDE11A, in part due to the lack of selective PDE11 inhibitors. However, recent reports with a PDE11 knockout mouse model have been interpreted to suggest that PDE11 may be important for sperm development and function, as well as psychiatric diseases such as schizophrenia (PNAS, 2010; 107(8); 8457-62), while gene association studies link mutations in PDE11A with adrenocortical tumors and Cushing's Syndrome in which cortisol levels in the blood are elevated. As such, a small molecule inhibitor of PDE11 could, and does, increase cortisol synthesis providing a therapeutic route to for treating adrenal insufficiency and adrenal insufficiency associated diseases. The present invention is directed toward overcoming one or more of the problems discussed above.